Wound healing compositions

ABSTRACT

Provided herein are biologically active solution compositions comprising one or more sacrificial proteolytic enzyme substrates, one or more preservatives, and one or more antimicrobial agents and methods of using the solution compositions to treat tissue sites, in particular chronic wounds. The compositions may be used in conjunction with negative pressure wound therapy to treat tissue sites.

RELATED APPLICATIONS

The present application is a Continuation of U.S. patent applicationSer. No. 14/206,914, entitled “WOUND HEALING COMPOSITIONS,” filed Mar.12, 2014, which claims the benefit, under 35 USC § 119(e), of the filingof U.S. Provisional Patent Application Ser. No. 61/799,367, entitled“WOUND HEALING COMPOSITIONS,” filed Mar. 15, 2013, which areincorporated herein by reference for all purposes.

FIELD

The present disclosure relates generally to compositions and methods formedical treatment of wounds, and more particularly but withoutlimitation, for treatment or prophylaxis of chronic wounds.

BACKGROUND

Typical procedures for treating chronic wounds such as, for example,venous ulcers, diabetic ulcers and pressure sores, include the use ofabsorbent dressings or hydrocolloid gels. Additionally, since mostchronic wounds are infected, many wound dressings contain antimicrobialagents, such as silver or iodine, to either create a barrier tomicroorganisms or reduce microbial load. These treatments are used morefor managing the wound environment and moisture balance than activelypromoting wound healing.

In wound healing, the extracellular matrix (ECM), comprised largely ofcollagen, plays a significant role in the healing response. Chronicwounds suffer from the fact that increased levels of inflammatory cellsand proteases are present and work to degrade the ECM, thereforeinhibiting its healing. Matrix metalloproteases (MMPs) are among theproteases present in both acute and chronic wounds and play an importantrole in the wound healing response. In normal wound healing, MMPs helpto degrade denatured ECM, which allows the functional matrix to beexposed. However, in chronic wounds, elevated numbers of MMPs and aresulting distortion in the ratio of MMPs to their inhibitors (tissueinhibitor metalloproteinase (TIMPs) cause disruption in the woundhealing system and can result in destruction of the ECM, growth factors,and granulation tissue.

The production of MMPs at a chronic wound site can be inhibited bypreventing activation of MMPs or by use of MMP inhibitors. A sacrificialsubstrate for the MMPs can also be employed to inhibit production. Somewound dressings on the market use various forms of natural collagen as asacrificial substrate for MMPs because the collagen also provides themechanical properties (integrity) necessary to form the dressing.However, with the loss of the collagen over time, the sacrificialsubstrate is no longer available. Accordingly, managing wounds and othertissue sites with elevated levels of MMPs continues to present asignificant challenge to healthcare providers and manufacturers.

SUMMARY

Described herein are bioactive compositions for treating tissue sites,in particular chronic wounds. The compositions described herein maycomprise one or more sacrificial proteolytic enzyme substrates and oneor more antimicrobial agents. The compositions may be solutions. As anillustrative embodiment, the solution compositions disclosed herein canconsist essentially of one or more MMP substrates, one or moreantimicrobial agents, and one or more preservatives such as sodiumbenzoate or chelators such as ethylenediaminetetraacetic acid (EDTA).

In one illustrative embodiment, the compositions are provided in adispensable liquid form, suited for instillation, for the management ofa tissue site through maintaining a moist wound bed along with the addedbenefit of MMP scavenging ability and antimicrobial effectiveness. Thisis an advancement over known methods and systems as a fresh batch of MMPsubstrate can be continually or cyclically delivered to a tissue siteduring each instillation.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 shows a block diagram of a representative embodiment of a therapysystem.

FIG. 2 shows the ability of an embodiment of the bioactive compositiondescribed herein to competitively inhibit MMP-9 activity at threedifferent enzyme protein concentrations.

FIGS. 3A and 3B show that the bioactive composition of FIG. 2 performssimilarly as a normal saline (0.9% NaCl) solution with a negativepressure wound therapy (NPWT) system. No degradation of the negativepressure was seen once the system stabilizes from the instill cycle seenby the initial peaks up to 1000 second time point.

DETAILED DESCRIPTION

In the following detailed description of the illustrative embodiments,reference is made to the accompanying drawings that form a part hereof.These embodiments are described in sufficient detail to enable thoseskilled in the art to practice the compositions and methods disclosedherein, and it is understood that other embodiments may be utilized andthat logical structural, mechanical, electrical, and chemical changesmay be made without departing from the spirit or scope of thedisclosure. To avoid detail not necessary to enable those skilled in theart to practice the embodiments described herein, the description mayomit certain information known to those skilled in the art. Moreover,descriptions of various alternatives using terms such as “or” do notnecessarily require mutual exclusivity unless clearly required by thecontext. The claimed subject matter may also encompass alternativeembodiments, variations, and equivalents not specifically described indetail. The following detailed description should therefore be taken asillustrative and not limiting.

As presented herein, a bioactive composition in the form of agelatin-based solution can competitively inhibit MMP activity, and thecomposition in the form of a solution can be used as an instillant tocleanse wounds. The combination of a MMP substrate and an antimicrobialagent in a dispensable liquid form, suited for instillation, enables themanagement of a tissue site through maintaining a moist wound bed inaddition to providing MMP scavenging ability and antimicrobialeffectiveness. To further promote healing and growth of tissue,embodiments of bioactive compositions including a preservative such as achelator, for example EDTA, and an antimicrobial agent, may worksynergistically to kill microbes while preventing further biofilmformation. This combination of the MMP substrate and antimicrobial agentcan create an optimal wound healing environment, allowing for mitigationof prolonged inflammation due mainly to excessive proteases present atthe tissue site and for progression into a normalized healing state.Also described herein is an innovative method of applying freshsacrificial MMP substrate to the tissue site, while moisturizing andcleansing the tissue with a fresh batch of antimicrobial agent at everycycle of instillation.

The term “tissue site” in this context broadly refers to a wound ordefect located on or within tissue, including but not limited to, bonetissue, adipose tissue, muscle tissue, neural tissue, dermal tissue,vascular tissue, connective tissue, cartilage, tendons, or ligaments. Awound may include, for example, chronic, acute, traumatic, sub-acute,and dehisced wounds, partial-thickness burns, ulcers (such as diabetic,pressure, or venous insufficiency ulcers), flaps, and grafts. The term“tissue site” may also refer to areas of any tissue that are notnecessarily wounded or defective, but are instead areas in which it maybe desirable to add or promote the growth of additional tissue. Forexample, reduced pressure may be used in certain tissue areas to growadditional tissue that may be harvested and transplanted to anothertissue location.

The term “topical” application refers to application to skin, dermis ortissue site, and application to such tissue sites may includeapplication adjacent to or within the tissue site.

The term “bioactive composition” or “biologically active composition” asused herein refers to a composition formulated with at least onesacrificial proteolytic enzyme substrate and at least one antimicrobialagent. Such compositions may be formulated in any carrier orpharmaceutically acceptable carrier and will typically comprise aneffective amount of sacrificial proteolytic enzyme substrate andantimicrobial agent to reduce inflammation and stimulate tissue healing.

The compositions disclosed herein are bioactive or biologicalcompositions comprising or consisting essentially of one or moresacrificial proteolytic enzyme substrates, one or more preservatives,and one or more antimicrobial agents. The term “consisting essentiallyof” as used herein functions to limit the scope to the specifiedmaterials or steps as well as those that do not materially affect thebasic and novel characteristic(s) of the claimed compositions ormethods. In one embodiment, the compositions comprise or consistessentially of a MMP substrate as the sacrificial proteolytic enzymesubstrate, EDTA as one of the preservatives, sodium benzoate as a secondpreservative, and one or more antimicrobial agents. The compositions mayinclude one or more carriers and/or other inert agents that do notmaterially affect the basic and novel characteristics of thecomposition.

The term “carrier” as used herein refers to diluents, adjuvants,excipients, vehicles, and other inert agents with which the sacrificialproteolytic enzyme substrate is administered.

Described herein are compositions consisting of one or more sacrificialproteolytic enzyme substrate, one or more preservatives, and one or moreantimicrobial agents. As an example, the composition consists of a MMPsubstrate as the sacrificial proteolytic enzyme substrate, EDTA andsodium benzoate as the preservatives, and one or more antimicrobialagents.

The compositions provided herein also include pharmaceuticalcompositions. The pharmaceutical compositions may include one or morepharmaceutically acceptable carriers and/or other pharmaceuticallyacceptable inert agents. The term “pharmaceutically acceptable” as usedherein refers to ingredients, agents, or compositions that are suitablefor pharmaceutical administration without undue toxicity,incompatibility, instability, irritation, allergic response and thelike. A “pharmaceutically acceptable salt” can be derived frompharmaceutically acceptable inorganic or organic bases and frompharmaceutically acceptable inorganic and organic acids, which are knownin the art and can be derived by one of ordinary skill in the art.Examples of pharmaceutically acceptable carriers include but are notlimited to sugars, starches, cellulose, excipients, oils, glycols,polyols, esters, agar, and buffering agents. The above are non-limitingexamples of carriers. Pharmaceutically acceptable carriers may bedistinct from carrier materials described below and are known in theart. Pharmaceutically acceptable carriers other than those listed hereinmay be easily formulated by those of ordinary skill in the art.

The compositions disclosed herein may additionally comprise conventionaladjuvants such as propionic acid, propylene glycol, conventionalbuffers, preservatives, hydrophilic emulsifiers, lipophilic emulsifiers,perfumes, emollients, deodorants, humectants and the like. Colorants mayalso optionally be added in the compositions disclosed herein. Adjuvantswhich would be harmful to a tissue site or surrounding skin should beavoided, as well as those adjuvants which may react with and/oradversely reduce the effectiveness of the composition.

The compositions disclosed herein may be formulated into a wide varietyof articles to be topically applied that include but are not limited tolotions, creams, gels, sticks, sprays, ointments, pastes, foams, powdersand film-forming products. The compositions provided herein may beformulated for debriding, irrigating, moisturizing, cleansing,lubricating, and/or disinfecting a tissue site. The composition may be aliquid formulation or in the form of a solution. Such compositions maybe formulated for time-controlled release.

The compositions may be an instillation composition. The compositionsmay be delivered to a tissue site by continuous instillation and/orperiodic instillation. The instillant provides fresh sacrificialproteolytic enzyme substrate, such as MMP substrate, and antimicrobialagent to the tissue site.

The compositions provided herein comprise sacrificial proteolytic enzymesubstrates for preventing activation of an enzyme. As discussed above,normal endogenous levels of MMPs are essential for tissue remodelingduring the healing process. However, in excess, they continually breakdown the new tissue that is formed. This leads to a wound that eitherdoes not heal quickly or becomes stalled. Excess levels of MMPs create asustained state of inflammation thereby preventing the progression ofnormal wound healing. Accordingly, in certain aspects, methods aredescribed herein for promoting healing or growth of tissue, comprisingproviding a composition containing a MMP substrate in an amounteffective to reduce the level of MMPs and/or reduce inflammation at atissue site and in surrounding tissue. Excessive MMP activity at atissue site can also be addressed by providing a composition comprisinga sacrificial proteolytic enzyme substrate, such as protein, proteinhydrolysate, or combinations thereof.

The compositions provided herein comprise one or more substrates for MMPas the sacrificial proteolytic enzyme substrate. Examples of MMPsubstrates include, but are not limited to, collagen, gelatin, elastin,casein, albumin, fibrinogen, fibronectin, and combinations andhydrolysates thereof. In certain embodiments, proteins for use assacrificial substrates are hydrolyzed or partially hydrolyzed bytreatment with a strong acid or base. Such treatment can fragment thesubject proteins and generate a more accessible peptide sequence to bindto proteolytic enzymes.

The compositions disclosed herein comprise about 0.01% to 25%, 0.01% to10%, 0.03% to 1%, 0.03% to 5%, or 1% to 10% w/v, or about 6%, 8%, 10%,15% or 20% w/v of sacrificial proteolytic enzyme substrate.

The most prevalent MMPs in chronic wounds are the gelatinase proteases,MMP-2 and MMP-9 that more readily target the hydrolyzed or denaturedform of collagen known as “gelatin.” Thus, in certain aspects, abioactive composition for use as described here further comprises acollagen, such as a hydrolyzed collagen (e.g., gelatin). Gelatin can beprocessed from a variety of sources including, but not limited to,bovine skin, porcine skin and bone material. Depending on the hydrolysismethods employed in manufacture, the gelatin may be defined as a type Aor type B gelatin. One advantage of using a gelatin rather than, or inaddition to, collagen is that gelatin includes exposed peptide sequencesthat serve as signals for protease binding. Accessibility of signalingsequences in the native collagen molecule is diminished due to thetriple-helix structure of the native collagen molecule, wherepolypeptide chains are bound with strong hydrogen bonds. Thus, incertain aspects, a composition is defined as not comprising collagen. Inthe case of gelatin, on the other hand, signaling sequences are readilyexposed to proteases making it more efficient as a sacrificialsubstrate.

A primary constraint against using gelatin in wound dressings has beeninsufficient mechanical integrity and inability to maintain dressingshape in the wound environment as is possible with natural collagen.However, if gelatin is applied as a coating onto another porousmaterial, such as a bandage, gauze, or polyurethane foam, which willprovide structural support, such material with gelatin may be anexcellent choice as a MMP sacrificial substrate. Therefore, in oneembodiment, gelatin for use in the bioactive solution compositionsprovided herein can comprise a molecular weight of between about 2000 Dato about 20,000 Da or having a bloom value of less than about 150. Incertain embodiments, it may be beneficial to use gelatin with sufficientgel strength (for example, a gelatin having a molecular weight of about2000 Da to about 20,000 Da or having a bloom value of less than about150) to form an adherent layer on a porous material without causing thematerial to become overly stiff.

Additionally, gelatin is an excellent oxygen barrier, which is importantfor stability of molecules that could be incorporated in wound dressingsfor instance, such as antioxidants and oxygen-sensitive proteins andpeptides. Thus, the carrier material within a wound dressing may be apolyurethane foam as described herein that is coated with gelatin toprovide the reduced pressure dressing with a sacrificial substrate forMMPs. The compositions may comprise, for example, 0.01% to 25%, 0.01% to10%, 0.03% to 1%, 0.03% to 5%, or 0.5% to 10% w/v, or about 1%, 2%, 3%,4% or 5% w/v of gelatin. The compositions may also comprise, forexample, 0.1% to 25%, 1% to 10% or about 6%, 8%, 10%, 15%, or 20% w/wgelatin.

The compositions provided herein may include one or more preservatives.The compositions may contain preservatives for the MMP substrate.Examples of preservatives include, but are not limited to chelators suchas EDTA, diethylene triamine pentaacetic acid (DTPA), and catechins;sodium benzoate; potassium sorbate; and sodium nitrate. EDTA is alsocapable of reducing MMP, and sodium benzoate works synergistically withEDTA. The compositions may comprise about 0.01% to about 5%, 0.1% to 3%,0.015% to 1%, 0.015% to 0.5%, 0.01% to 0.1%, or 0.0225% to 0.1% w/v orabout 0.015%, 0.225%, or 0.1% w/v.

The compositions provided herein comprise one or more antimicrobialagents. The antimicrobial agents can act to counter any bacterialprotease activity that may hamper the healing environment, which allowsa tissue site to progress towards an optimal healing state. Examples ofantimicrobial agents include, but are not limited to, components of aloevera, ashitaba, bacteriophage, beta-defensin, quaternary ammoniumcompound, chlorhexidine, copper, dispersin B, essential oil, gentamicin,lactoferrin, lysostaphin, N-halamines, nitric oxide, oleic acid, PLUNC,polyhexanide biguanide (PHMB), bacteriocin, selenium, silver compound,triclosan, zinc, and combinations thereof. Aloe vera contains numerousphotochemical compounds including but not limited to tannin, saponin,flavonoids, and fumaric acid. As used herein, the term “PLUNC” refers tothe gene or clone encoding the palate, lung, nasal epithelium carcinomaassociated protein and to the protein itself. Examples of quaternaryammonium compound include benzethonium chloride and benzalkoniumchloride. An example of a beta-defensin is cathelicidin (LL-37).Examples of a silver compound may include colloidal silver, ionicsilver, nonionic silver, silver chloride, silver nanopartices, andsilver sulfadiazine. Examples of essential oil include but are notlimited to cinnamon oil, clove oil, eucalyptus oil, and tea tree oil. Anexample of chlorhexidine is chlorhexidine gluconate. The compositionsmay comprise about 0.01% to 1%, 0.05% to 1%, or 0.05% to 0.5% w/v ofantimicrobial agents.

The compositions disclosed herein may further comprise other agents suchas growth factors, cytokines, and proteinase inhibitors, in particularproteinase inhibitors of MMPs. The compositions provided herein mayconsist essentially of or consist of one or more MMP substrates, one ormore preservatives, one or more antimicrobial agents, one or more growthfactors, and one or more proteinase inhibitors.

In certain aspects, the composition is sterilized by irradiation. Askilled worker will recognize that such irradiation can alter the amountof cross-linking within proteins in the composition. Thus, in caseswhere the composition comprises a sacrificial proteolytic enzymesubstrate, such as a MMP substrate, that is a protein, such as gelatin,the amount of irradiation may be limited to prevent proteincross-linking, while still achieving sterilization.

In one embodiment, application of the composition provided herein may beinfusion within, injection into, absorption by, layering on,encapsulation within, or coating on, a carrier material, such as abandage, gauze, wound dressing, adhesive bandage, scaffold, or hydrogel.A “carrier material” as used herein refers to a material suitable forhaving a proteolytic enzyme substrate, such as a MMP substrate and anantimicrobial agent. For example, a composition of a proteolytic enzymesubstrate and an antimicrobial agent may be applied to a woven,non-woven, or knitted fabric material, such as gauze, dispersed withinfilm, sponge, or foam for sustained release at a tissue site. Thecarrier material may be either bioresorbable, for instance comprisingpolyglycolic acid, polylactic acid, polydioxanone, polyhydroxybutyrate,polyhydrozyvalerate, polyaminoacids polyorthoesters, polyvinly alcohol,collagen, gelatin, chitosan, oxidized regenerated cellulose, hyaluronicacid, alginate or derivatives thereof, or may be non-bioresorbable,comprising for instance, polyurethane, polyvinyl alcohol, or gauze. Insome embodiments, the carrier material may be made of the same substanceas the proteolytic enzyme substrate, for instance collagen or a modifiedcollagen, such as gelatin. Carrier materials are distinct from thecarriers and pharmaceutically acceptable carriers used in bioactivecompositions.

Suitable carrier materials include, but are not limited to: bandages,gauze, wound dressings, adhesive bandages, scaffold, hydrogelscontaining cellulose derivatives, including hydroxyethyl cellulose,hydroxymethyl cellulose, carboxymethyl cellulose, hydroxypropylmethylcellulose and mixtures thereof; and hydrogels containing polyacrylicacid (Carbopols) as well as gelatin. The above carrier materials mayinclude alginate (as a thickener or stimulant), buffers to control pHsuch as disodium hydrogen phosphate/sodium dihydrogen phosphate, agentsto adjust osmolarity such as sodium chloride, and stabilizers such asEDTA.

The compositions provided herein are useful for the treatment of atissue site by any method where the composition contacts the tissuesite. For instance, the composition may contact a tissue site throughdirect application of a cream, a gel, an ointment or a spray. In anotherembodiment, the composition may be applied to a carrier material, whichis then applied to a tissue site. Such methods may include applicationof the composition to a bandage, gauze, or dressing to be applied to atissue site. The compositions provided herein may also be added to otherknown compositions for treating wounds or other tissue sites.

Also described herein are methods of using the disclosed composition fortreating a tissue site by debriding, irrigating, moisturizing,cleansing, lubricating, and/or disinfecting the tissue site. Thecomposition can deliver agents, including MMP substrates,antimicrobials, or growth factors, for example, to a tissue site, andcan deliver such agents in a manner and/or sequence to debride eschar,necrotic tissue, and debris; cleanse, irrigate, moisturize, disinfect,and remove/reduce wound bioburden and microbial biofilms; retardmicrobial and biofilm regrowth; decrease pain, odor, inflammation; andpromote wound healing physiology. The composition delivered to a tissuesite can provide a moist environment to promote healing.

For example, the bioactive compositions presented herein may beintegrated with negative pressure wound therapy (NPWT), fluidinstillation therapy, or both. Clinical studies and practice have shownthat NPWT can augment and accelerate growth of new tissue at a tissuesite. The applications of this phenomenon are numerous, but it hasproven particularly advantageous for treating wounds. Regardless of theetiology of a wound, whether trauma, surgery, or another cause, propercare of the wound is important to the outcome. Treatment of wounds withreduced pressure may be commonly referred to as NPWT, but is also knownby other names, including “negative-pressure therapy,” “reduced-pressurewound therapy,” “vacuum therapy,” and “vacuum-assisted closure,” forexample. Negative-pressure therapy may provide a number of benefits,including migration of epithelial and subcutaneous tissues, improvedblood flow, and micro-deformation of tissue at a tissue site. Together,these benefits can increase development of granulation tissue and reducehealing times.

Instillation of a tissue site, which generally refers to the slowintroduction of a solution to the tissue site, can expose a tissue siteto temperature variations, drugs, or other substances that may furtherpromote healing or growth of tissue. Instillation may also be referredto as irrigation or infusion in some contexts. Instillation may becontinuous or intermittent and may take place prior to, subsequent to,or simultaneously with the application of negative pressure. In someembodiments, instillation and negative pressure may be coordinated by acentral controller.

FIG. 1 is a simplified functional block diagram of an example embodimentof a therapy system 100 that can provide therapeutic pressure andinstillation in accordance with this specification. As illustrated, thetherapy system 100 may include a dressing 102 fluidly coupled to anegative-pressure source 104. A regulator or controller, such asregulator 106, may also be fluidly coupled to the dressing 102 and thenegative-pressure source 104. The dressing 102 generally includes adrape, such as drape 108, and a manifold, such as distribution manifold110. The therapy system 100 may also include fluid containers, such ascontainer 112 and container 114, coupled to the dressing 102. Asillustrated in FIG. 1, container 112 may be also be fluidly coupled tothe negative-pressure source 104 in some embodiments, and container 114may be coupled to a fluid-delivery device, such as a pump 116.

In general, components of the therapy system 100 may be coupled directlyor indirectly. For example, the negative-pressure source 104 may bedirectly coupled to regulator 106 and indirectly coupled to dressing 102through regulator 106. Components may be fluidly coupled to each otherto provide a path for transferring fluids (i.e., liquid and/or gas)between the components. In some embodiments, components may be fluidlycoupled with a tube, for example. A “tube,” as used herein, broadlyrefers to a tube, pipe, hose, conduit, or other structure with one ormore lumens adapted to convey fluids between two ends. Typically, a tubeis an elongated, cylindrical structure with some flexibility, but thegeometry and rigidity may vary. In some embodiments, components mayadditionally or alternatively be coupled by virtue of physicalproximity, being integral to a single structure, or being formed fromthe same piece of material. Coupling may also include mechanical,thermal, electrical, or chemical coupling (such as a chemical bond) insome contexts.

In operation, the distribution manifold 110 may be placed within, over,on, or otherwise proximate to a tissue site. The drape 108 may be placedover the distribution manifold 110 and sealed to tissue proximate to thetissue site. The tissue proximate to the tissue site is often undamagedepidermis peripheral to the tissue site. Thus, the dressing 102 canprovide a sealed therapeutic environment proximate to a tissue site,substantially isolated from the external, ambient environment. Thenegative-pressure source 104 can reduce the pressure in the sealedtherapeutic environment, and the pump 116 can apply therapeuticsolutions, including the embodiments of the bioactive compositionsdescribed herein. Reduced pressure and/or fluids can be appliedsubstantially uniformly through the distribution manifold 110 in thesealed therapeutic environment. Reduced pressure can induce macrostrainand microstrain in the tissue site, as well as remove exudate and otherfluids from the tissue site, which can be collected in the container 112and disposed of properly.

Integrating negative pressure therapy and instillation therapy withembodiments of bioactive compositions described herein can furtherpromote healing and growth of tissue by removing barriers to normalhealing, such as abnormally high levels of MMPs. To further promotehealing and growth of tissue, embodiments of bioactive compositionsincluding a preservative such as a chelator like EDTA and anantimicrobial agent, may work synergistically to kill microbes whilepreventing further biofilm formation. Functionally coupling infusion ofthe compositions with NPWT as disclosed herein provides unexpecteddecreases in wound bioburden and wound healing trajectories. The abilityof a gelatin-based solution to operate with a NPWT system allows for theuse of the solution as an instillate to cleanse a tissue site, inparticular a chronic wound, with bioactive and antimicrobial agents.

In some embodiments, the negative pressure with the bioactivecomposition can be applied during debridement of a tissue site.Alternatively, negative pressure therapy may be applied afterdebridement, to promote vascular stimulation and the formation ofgranulation tissue. Further still, the transition from debridement tonegative pressure therapy is seamless, as well as from negative pressuretherapy to passive infusion with the composition, that is, withoutdisrupting the integrity of the tissue site.

The negative pressure with the bioactive composition may also be appliedduring cleansing or irrigation of the wound in some embodiments.Alternatively, the negative pressure may be applied prior to or afterthe cleansing of the wound with the composition.

The compositions provided herein can be used in conjunction with allcurrent NPWT devices, and delivered in either the inpatient oroutpatient setting. Exemplary negative pressure devices include V.A.C.®Therapy, V.A.C. Instill®, or V.A.C. Ulta® therapy systems (KineticConcepts, Inc.). These devices or devices having similar or equivalentdesigns may be used.

“Negative pressure” or “reduced pressure” generally refers to a pressureless than a local ambient pressure, such as the ambient pressure in alocal environment external to a sealed therapeutic environment providedby the dressing 102. In many cases, the local ambient pressure may alsobe the atmospheric pressure in the vicinity of a tissue site.Alternatively, the pressure may be less than a hydrostatic pressureassociated with tissue at the tissue site. Unless otherwise indicated,values of pressure stated herein are gauge pressures. Similarly,references to increases in reduced pressure typically refer to adecrease in absolute pressure, while decreases in reduced pressuretypically refer to an increase in absolute pressure.

A negative-pressure source, such as the negative-pressure source 104,may be a reservoir of air maintained at a negative pressure, or may be amanual or electrically-powered device that can reduce the pressure in asealed volume, such as a vacuum pump, a suction pump, a wall suctionport available at many healthcare facilities, or a micro-pump, forexample. A negative-pressure source may be housed within or used inconjunction with other components, such as sensors, processing units,alarm indicators, memory, databases, software, display devices, or userinterfaces that further facilitate negative-pressure therapy. While theamount and nature of negative pressure applied to a tissue site may varyaccording to therapeutic requirements, the pressure typically rangesbetween −5 mm Hg (−667 Pa) and −500 mm Hg (−66.7 kPa). Commontherapeutic ranges are between −75 mm Hg (−9.9 kPa) and −300 mm Hg(−39.9 kPa).

A fluid-delivery device, such as the pump 116, may be a rotary-deliverypump, or other pump that can supply an instillation solution to a sealedspace or the distribution manifold 110. A fluid-delivery device may behoused within a therapy device or used in conjunction with othercomponents, such as sensors, processing units, alarm indicators, memory,databases, software, display devices, or user interfaces that furtherfacilitate instillation therapy. In some embodiments, a fluid-deliverydevice and a negative-pressure source may be integrated into a singleunit to provide both negative pressure and instillation, or toalternatingly supply negative pressure and instillation.

A manifold, such as the distribution manifold 110, can generally beadapted to contact a tissue site. The distribution manifold 110 may beadapted to be placed partially or fully in contact with the tissue site.If the tissue site is a wound, for example, the distribution manifold110 may partially or completely fill the wound, or may be placed overthe wound. The distribution manifold 110 may take many forms, and may bemany sizes, shapes, or thicknesses depending on a variety of factors,such as the type of treatment being implemented or the nature and sizeof a tissue site. For example, the size and shape of the distributionmanifold 110 may be adapted to the contours of deep and irregular shapedtissue sites.

More generally, a manifold is a substance or structure adapted todistribute negative pressure to or remove fluids from a tissue site, orboth. In some embodiments, though, a manifold may also facilitatedelivering fluids to a tissue site, if the fluid path is reversed or asecondary fluid path is provided, for example when instillation solutionis applied. A manifold may include flow channels or pathways thatdistribute fluids provided to and removed from a tissue site around themanifold. In one illustrative embodiment, the flow channels or pathwaysmay be interconnected to improve distribution of fluids provided to orremoved from a tissue site. For example, cellular foam, open-cell foam,porous tissue collections, and other porous material such as gauze orfelted mat generally include structural elements arranged to form flowchannels. Liquids, gels, and other foams may also include or be cured toinclude flow channels.

In one illustrative embodiment, the distribution manifold 110 may be aporous foam material having interconnected cells or pores adapted touniformly (or quasi-uniformly) distribute reduced pressure to a tissuesite. The foam material may be either hydrophobic or hydrophilic. In onenon-limiting example, the distribution manifold 110 may be an open-cell,reticulated polyurethane foam such as GranuFoam® dressing available fromKinetic Concepts, Inc. of San Antonio, Tex.

In some embodiments, such as embodiments in which the distributionmanifold 110 may be made from a hydrophilic material, the distributionmanifold 110 may also wick fluid away from a tissue site whilecontinuing to distribute reduced pressure to the tissue site. Thewicking properties of the distribution manifold 110 may draw fluid awayfrom a tissue site by capillary flow or other wicking mechanisms. Anexample of a hydrophilic foam is a polyvinyl alcohol, open-cell foamsuch as V.A.C. WhiteFoam® dressing available from Kinetic Concepts, Inc.of San Antonio, Tex. Other hydrophilic foams may include those made frompolyether. Other foams that may exhibit hydrophilic characteristicsinclude hydrophobic foams that have been treated or coated to providehydrophilicity.

The distribution manifold 110 may further promote granulation at atissue site if pressure within a sealed therapeutic environment isreduced. For example, any or all of the surfaces of the distributionmanifold 110 may have an uneven, coarse, or jagged profile that caninduce microstrains and stresses at a tissue site if reduced pressure isapplied through the distribution manifold 110.

In one example embodiment, the distribution manifold 110 may beconstructed from bioresorbable materials. Suitable bioresorbablematerials may include, without limitation, a polymeric blend ofpolylactic acid (PLA) and polyglycolic acid (PGA). The polymeric blendmay also include without limitation polycarbonates, polyfumarates, andcapralactones.

Other bioresorbable materials that may be used include, but are notlimited to, polydioxanone, polyhydroxybutyrate, polyhydrozyvalerate,polyaminoacids polyorthoesters, polyvinly alcohol, chitosan, oxidizedregenerated cellulose, hyaluronic acid, alginate, collagen, a modifiedcollagen, such as gelatin or derivatives of any of the above.

The distribution manifold 110 may further serve as a scaffold for newcell-growth, or a scaffold material may be used in conjunction with thedistribution manifold 110 to promote cell-growth. In general, a scaffoldmaterial may be a substance or structure used to enhance or promote thegrowth of cells or formation of tissue, such as a three-dimensionalporous structure that provides a template for cell growth.

A scaffold and/or manifold may be also be infused with, coated with, orcomprised of cells, growth factors, extracellular matrix components,nutrients, integrins, or other substances to promote cell growth inaddition to embodiments of the compositions described herein. Themanifold or scaffold may serve as a carrier material for the compositiondescribed herein.

Scaffolds may be formed from biologic or synthetic scaffold materials,and are used in the field of tissue engineering to support proteinadhesion and cellular ingrowth for tissue repair and regeneration. Thecurrent state of the art in scaffold technology relies upon the inherentcharacteristics of the surrounding tissue space for the adsorption ofproteins and migration of cells. Nonlimiting examples of suitablescaffold materials include extracellular matrix proteins such as fibrin,collagen or fibronectin, and synthetic or naturally occurring polymers,including bioabsorbable or non-absorbable polymers, such as polylacticacid (PLA), polyglycolic acid (PGA), polylactide-co-glycolide (PLGA),polyvinylpyrrolidone, polycaprolactone, polycarbonates, polyfumarates,caprolactones, polyamides, polysaccharides (including alginates (e.g.,calcium alginate) and chitosan), hyaluronic acid, polyhydroxybutyrate,polyhydroxyvalerate, polydioxanone, polyorthoesthers, polyethyleneglycols, poloxamers, polyphosphazenes, polyanhydrides, polyamino acids,polyacetals, polycyanoacrylates, polyurethanes (e.g., GranuFoam®),polyacrylates, ethylene-vinyl acetate polymers and other acylsubstituted cellulose acetates and derivatives thereof, polystyrenes,polyvinyl chloride, polyvinyl fluoride, poly(vinylimidazole),chlorosulphonated polyolefins, polyethylene oxide, polyvinyl alcohol,Teflon®, and nylon.

The scaffold can also comprise ceramics such as hydroxyapatite,coralline apatite, calcium phosphate, calcium sulfate, calcium carbonateor other carbonates, bioglass, allografts, autografts, xenografts,decellularized tissues, or composites of any of the above. In someembodiments, the scaffold may comprise collagen (e.g., Biostep® orPromogran® scaffolds), polylactic acid (PLA), polyglycolic acid (PGA),polylactide-co-glycolide (PLGA), a polyurethane, a polysaccharide, anhydroxyapatite, or a polytherylene glycol. Additionally, the scaffoldcan comprise combinations of any two, three or more materials, either inseparate or multiple areas of the scaffold, combined noncovalently orcovalently (e.g., copolymers such as a polyethylene oxide-polypropyleneglycol block copolymers, or terpolymers), or combinations thereof.

The drape 108 is an example of a sealing member. A sealing member may beconstructed from a material that can provide a fluid seal between twoenvironments or components, such as between a therapeutic environmentand a local external environment. The sealing member may be, forexample, an impermeable or semi-permeable, elastomeric material that canprovide a seal adequate to maintain a negative pressure at a tissue sitefor a given negative-pressure source. For semi-permeable materials, thepermeability generally should be low enough that a desired negativepressure may be maintained. An attachment device may be used to attach asealing member to an attachment surface, such as undamaged epidermis, agasket, or another sealing member. The attachment device may take manyforms. For example, an attachment device may be a medically-acceptable,pressure-sensitive adhesive that extends about a periphery, a portion,or an entire sealing member. Other example embodiments of an attachmentdevice may include a double-sided tape, paste, hydrocolloid, hydrogel,silicone gel, organogel, or an acrylic adhesive.

The container 112 is representative of a container, canister, pouch, orother storage component, which can be used to manage exudates and otherfluids withdrawn from a tissue site. In many environments, a rigidcontainer may be preferred or required for collecting, storing, anddisposing of fluids. In other environments, fluids may be properlydisposed of without rigid container storage, and a re-usable containercould reduce waste and costs associated with negative-pressure therapy.

The container 114 is representative of another container, canister,pouch, cartridge, or other storage component, which can be used tomanage instillation solution to be supplied to a tissue site. In manyenvironments a rigid container may be preferred or required fordelivering, storing, and supplying of the instillation solution. Inother environments, instillation solution may be provided in a non-rigidcontainer, and a re-usable container could reduce waste and costsassociated with instillation.

Components of therapy system 100 may also be provides as one or morekits. In one embodiment, for example, a kit comprises the systemdescribed above and one or more embodiments of a bioactive compositiondescribed herein. In another embodiment, a kit comprises one or moreembodiments of a bioactive composition described herein and an apparatusfor delivering the composition to a tissue site. The apparatus mayinclude a dressing.

The systems and methods described herein may provide significantadvantages, some of which have already been mentioned. For example, thecompositions, apparatuses, methods, systems, and kits described hereincan enable the delivery of agents to tissue sites that may not have beenreachable with a conventional collagen dressing, since a liquid basedsolution follows a path of least resistance. Moreover, moisture and MMPscavenging can be maintained at a tissue site. The added antimicrobialactivity to the compositions described herein can also provide bacterialkilling ability and reduce proteolytic activity. The viscosity and theadhesive nature of the gelatin enhance these effects.

EXAMPLES

The following examples are included to demonstrate the advantages andunexpected results of certain embodiments. Those of skill in the artshould, in light of the present disclosure, appreciate that many changescan be made to the specific embodiments which are disclosed and stillobtain a like or similar result without departing from the scope of theclaims. More specifically, it will be apparent that certain agents,which are both chemically and physiologically related, may besubstituted for the agents described herein while the same or similarresults would be achieved. All such similar substitutes andmodifications apparent to those skilled in the art are deemed to bewithin the scope of the appended claims.

Example 1: Gelatin Instillant Wound Cleanse Solution

Gelatin solution was prepared by dissolving gelatin in water to make a50 mg/mL stock solution. Serial dilutions were made by mixing the stocksolution with additional water. A stock MMP-9 proenzyme was activatedwith APMA per a method suggested by the enzyme manufacturer. Activatedproenzyme is diluted to the appropriate concentration in assay buffer.MMP-9 and gelatin solutions are added in specific quantities (determinedby desired concentration of each) to each assay well.

MMP-9 activity is measured by reading a colorimetric reaction whichresults from hydrolysis of a thioester bond by active MMP-9 resulting inproduction of a sulfhydryl group which then reacts with Ellman'sreagent. Higher absorbance readings correlate to higher MMP-9 activity.

FIG. 2 shows the ability of gelatin solution to competitively inhibitMMP activity at three different enzyme protein concentrations.

Example 2: Performance of Gelatin Solution with NPWTi

The V.A.C. Veraflo® dressing was placed on a sheet of 0.5 inch thickacrylic approximately 3 inch×3 inch square shaped and covered utilizingV.A.C. Veraflo® advanced drape. Therapy was set to Instill withinstillation volume set at 35 cc with a soak time of 10 minutes. Therapytime was 3.5 Hours at −125 mmHg. Data points were then read across a36-manometer pad and averaged as seen in FIG. 3.

The gelatin solution performs with the V.A.C. ULTA®, a NPWTi system fromKinetic Concepts, Inc., with high similarity to a normal salinesolution. FIG. 3A and FIG. 3B show the distribution of pressure with thegelatin solution and with normal saline. As shown, there is nodegradation of negative pressure after the system stabilizes from theinstill cycle. FIGS. 3A and 3B only show initial peaks up to the 1000second time point as the system stabilizes.

Although the present invention and its advantages have been disclosed inthe context of certain illustrative, non-limiting embodiments, it shouldbe understood that various changes, substitutions, permutations, andalterations can be made without departing from the scope of theinvention as defined by the appended claims.

We claim:
 1. A pharmaceutical composition for use in a method of woundhealing, wherein the pharmaceutical composition is configured to beadministered to a tissue site by fluid instillation therapy in the formof a solution, wherein the composition comprises: one or more matrixmetalloprotease (MMP) substrates comprising gelatin or a hydrolysatethereof; one or more preservatives selected from the group consisting ofethylenediaminetetraacetic acid (EDTA), diethylene triamine pentaaceticacid (DTPA), catechins, sodium benzoate, potassium sorbate, and sodiumnitrate; one or more antimicrobial agents; and a pharmaceuticallyacceptable carrier.
 2. The pharmaceutical composition of claim 1,wherein the antimicrobial agent is selected from the group consisting ofaloe vera components, ashitaba, bacteriophage, beta-defensin, quaternaryammonium compound, chlorhexidine, copper, dispersin B, essential oil,gentamicin, lactoferrin, lysostaphin N-halamines, nitric oxide, oleicacid, PLUNC protein, polyhexanide biguanide (PHIVIB), bacteriocin,selenium, silver compound, triclosan, zinc, and combinations thereof. 3.The pharmaceutical composition of claim 2, wherein the quaternaryammonium compound is benzethonium chloride or benzalkonium chloride. 4.The pharmaceutical composition of claim 2, wherein the beta-defensin iscathelicidin (LL-37).
 5. The pharmaceutical composition of claim 2,wherein the silver compound is colloidal silver, ionic silver, silverchloride, silver nanopartices, or silver sulfadiazine.
 6. Thepharmaceutical composition of claim 2, wherein the essential oil iscinnamon oil, clove oil, eucalyptus oil, or tea tree oil.
 7. Thepharmaceutical composition of claim 2, wherein the chlorhexidine ischlorhexidine gluconate.
 8. The pharmaceutical composition of claim 1,further comprising one or more growth factors.
 9. The pharmaceuticalcomposition of claim 1, further comprising one or more proteinaseinhibitors that are proteinase inhibitors of MMPs.
 10. Thepharmaceutical composition of claim 1, wherein the gelatin comprises amolecular weight of between 2000 Da to 20,000 Da.
 11. The pharmaceuticalcomposition of claim 1, wherein the gelatin has a bloom value of lessthan
 150. 12. The pharmaceutical composition of claim 1, wherein thepharmaceutical composition comprises 0.1% to 25% w/w gelatin.
 13. Thepharmaceutical composition of claim 1, wherein the pharmaceuticalcomposition comprises 1% to 10% w/w gelatin.
 14. A kit comprising acontainer and a composition within the container, the compositioncomprising: one or more matrix metalloprotease (MMP) substrates, whereinthe MMP substrates comprise gelatin or a hydrolysate thereof, one ormore preservatives comprising a chelating agent, wherein the chelatingagent is ethylenediaminetetraacetic acid (EDTA), and apharmaceutically-acceptable carrier; wherein the composition is providedin a dispensable liquid form for instillation to a tissue site.
 15. Thekit of claim 14, further comprising a dressing having a first portionadapted for contact with the tissue site and a second portion in fluidcommunication with the container.
 16. The kit of claim 15, wherein thedressing comprises an open-cell reticulated polyurethane foam pad. 17.The kit of claim 16, wherein the foam pad is treated with thecomposition prior to use.
 18. The kit of claim 16, wherein the foam padis infused and coated on the surface with the composition.
 19. The kitof claim 14, wherein the composition further comprises one or moreantimicrobial agents.
 20. The kit of claim 14, wherein the compositionfurther comprises one or more growth factors.